Biochemical experiments proposed in this application seek to identify features of nucleic acid sequence and structure that function specifically as substrates for RNA polymerase and RNA processing enzymes, that trigger the termination step during RNA synthesis, and that distinguish efficient from inefficient initiator regions for protein biosynthesis. Experimental systems are chosen to permit both in vitro biochemical analysis and studies of the in vivo functioning of these elements. The functional determinants of signals for transcription termination, RNA processing, and ribosome binding will be identified by systematic manipulation of nucleotide sequence. Nucleases specific for both single and double strands and chemical reagents will be used as experimental probes of RNA secondary structure to obtain evidence for intramolecular interactions believed to strongly influence mRNA activity during steps of gene expression. The capacity for independent recognition by ribosomes of several initiators in in vitro and in vivo assays will be examined. Translational yields will be measured under conditions that provide a standard for reference.